pathogenic role in disease

Einleitung Vaskulopathie, entzündliche Fibrose und funktionelle Autoantikörper (Ak) sind Hauptmanifestationen der systemischen Sklerose, einer schwerwiegenden rheumatischen Erkrankung mit einer hohen Sterblichkeitsrate, begrenzten therapeutischen Möglichkeiten und ungeklärter Krankheitsursache. Wir haben Ak gerichtet gegen den Angiotensin II Typ 1 Rezeptor (AT1R) und den Endothelin-1 Typ A Rezeptor (ETAR) in Patienten mit systemischer Sklerose identifiziert. Klinische Untersuchungen zeigten Assoziationen dieser Ak mit charakteristischen Merkmalen der systemischen Sklerose. Wir stellen die Hypothese auf, dass diese Ak krankheitsverursachende Effekte auslösen können. Der Einfluss von anti-AT1R und anti-ETAR Ak zur Initialisierung von Entzündung und Fibrose wurde analysiert. Methodik Anti-AT1R und anti-ETAR Ak-positives Immunglobulin G (IgG) von Patienten mit systemischer Sklerose (SSc-IgG) wurde verwendet. IgG aus gesunden Spendern (NC-IgG) diente als Negativ-Kontrolle. Die Aktivierung von AT1R und ETAR wurde mittels Antagonisten inhibiert. Die Protein-Expression wurde mit ELISA und mRNA-Expression mittels Real Time-PCR gemessen. Die endotheliale Wundheilung wurde mittels einer Wundheilungs- Untersuchung und die Kollagen-Expression mittels Immunocytochemie bestimmt. Transendotheliale Migration von Neutrophilen wurde mittels Zellkultureinsätzen und Aktivierung von reaktiven Sauerstoffspezies mittels Immunfluoreszenz analysiert. Zelluläre Zusammensetzung und die Anzahl der Neutrophilen in bronchoalveolären Lavage-Fluiden (BALF) wurden mittels Durchlichtmikroskopie bestimmt nach passivem Transfer von SSc-IgG oder NC-IgG in naïve C57BL/6J Mäuse. Plasmaspiegel von KC (murines funktionelles Homolog zum humanen Interleukin-8) wurden mittels eines Suspensions-Test-System überprüft. Histologische Analysen wurden mittels Durchlichtmikroskopie durchgeführt. Ergebnisse Anti-AT1R und anti-ETAR Ak-positives SSc-IgG verursachte eine starke Aktivierung der humanen mikrovaskulären Endothelzellen (human microvascular endothelial cells-1, HMEC-1). Erhöhte Protein- und mRNA-Spiegel des pro-entzündlichen Chemokins IL-8 und erhöhte mRNA-Spiegel des vaskulären Zelladhäsionsmoleküls-1 wurden in HMEC-1 induziert. Darüber Abstrakt in deutscher Sprache hinaus, erhöhte die Aktivierung von HMEC-1 mit SSc-IgG die Transendotheliale Migration von Neutrophilen. Zudem zeigten Neutrophile, die mit Überständen von aktivierten HMEC-1 behandelt wurden, eine Aktivierung von reaktiven Sauerstoffspezies. SSc-IgG verminderte desweiteren die Wundheilung von HMEC-1 und verursachte unmittelbar eine Kollagenprotein-Typ I-Produktion in dermalen Fibroblasten. Effekte der Migration, Wundheilung und Kollagen- Produktion waren abhängig von Ak-Spiegeln. Passiver Transfer von anti-AT1R und anti-ETAR Ak-positiven SSc-IgG in naïve C57BL/6J Mäuse erhöhte die Neutrophilen-Anzahl in BALF. Parallel dazu, wurden erhöhte Spiegel von KC im Plasma von SSc-IgG-behandelten Mäusen gefunden ebenso wie strukturelle Veränderungen der Lungen. Schlussfolgerungen Wir kommen zum Schluss, dass die Aktivierung von Angiotensin- und Endothelin-Rezeptoren durch anti-AT1R und anti-ETAR Ak pathogene Effekte vermittelt und auf den Beitrag dieser Ak an der Pathogenese der SSc hinweist. Demzufolge können anti-AT1R und anti-ETAR Ak dabei helfen unser derzeitiges Verständnis dieser Erkrankung zu verbessern und dadurch neue Angriffsziele für therapeutische Intervention in der Behandlung von SSc zu erforschen.Introduction Vasculopathy, inflammatory fibrosis and functional autoantibodies (Abs) are major manifestations of systemic sclerosis (SSc) a severe rheumatic disease with high mortality rates, limited therapeutic options and unclear etiology. We have identified Abs directed against the angiotensin II type 1 receptor (AT1R) and endothelin-1 type A receptor (ETAR) in patients with SSc. Clinical analyses revealed an association with characteristic SSc features. We hypothesized that these Abs could facilitate pathogenic effects. The impact of anti-AT1R and anti-ETAR Abs on initiation of inflammation and fibrosis was analyzed. Methods Anti-AT1R and anti-ETAR Ab positive immunoglobulin G (IgG) from SSc patients (SSc-IgG) was used. Healthy donor IgG (NC-IgG) served as a normal control. AT1R and ETAR activation was inhibited by antagonists. Protein expression was measured by ELISA, mRNA expression by Real Time-PCR. Endothelial repair was measure by scratch assay and collagen expression by immunocytochemistry. Transendothelial migration of neutrophils was measured by a culture-insert-system and reactive oxygen species (ROS) activation by immunofluorescence. Cellular composition and neutrophil counts in bronchoalveolar lavage fluids (BALF) were analyzed microscopically after passive transfer of SSc-IgG or NC-IgG into naïve C57BL/6J mice. The plasma levels of KC (murine functional homologue to human interleukin-8), were quantified by a suspension array system. Histological analyses were performed using light microscopy. Results Anti-AT1R and anti-ETAR Ab positive SSc-IgG induced a strong activation of human microvascular endothelial cells (HMEC-1). Elevated protein and mRNA levels of the proinflammatory chemokine IL-8 and elevated mRNA levels of the vascular cell adhesion molecule-1 were induced in HMEC-1. Furthermore, activation of HMEC-1 with SSc-IgG increased the transendothelial neutrophil migration. Moreover, neutrophils exposed to supernatants of activated HMEC-1, showed an activation of ROS. SSc-IgG additionally decreased the wound repair of HMEC-1 and directly induced type I collagen production in fibroblasts. Effects of migration, wound repair and collagen expression were dependent on the Ab-levels. Passive transfer of anti- AT1R and anti-ETAR Ab positive SSc-IgG into naïve Abstrakt in englischer Sprache C57BL/6J mice increased neutrophil BALF counts. In parallel, increased levels of the chemokine KC, were found in the plasma of SSc-IgG treated mice as well as structural alterations of the lungs. Conclusions We conclude that angiotensin and endothelin receptor activation via anti-AT1R and anti-ETAR Abs mediate pathogenic effects, indicating their contribution to pathogenesis of SSc. Therefore, anti-AT1R and anti-ETAR Abs could help to improve our current understanding of this disease and thereby provide novel targets for therapeutic intervention in the treatment of SSc

Sleep–wake cycles and cognitive functioning in schizophrenia

Background Irregular sleep–wake cycles and cognitive impairment are frequently observed in schizophrenia, however, how they interact remains unclear. Aims To investigate the repercussions of circadian rhythm characteristics on cognitive performance and psychopathology in individuals with schizophrenia. Method Fourteen middle-aged individuals diagnosed with schizophrenia underwent continuous wrist actimetry monitoring in real-life settings for 3 weeks, and collected saliva samples to determine the onset of endogenous melatonin secretion as a circadian phase marker. Moreover, participants underwent multiple neuropsychological testing and clinical assessments throughout the study period. Results Sleep–wake cycles in individuals with schizophrenia ranged from well entrained to highly disturbed rhythms with fragmented sleep epochs, together with delayed melatonin onsets and higher levels of daytime sleepiness. Participants with a normal rest–activity cycle (objectively determined by high relative amplitude of day/night activity) performed significantly better in frontal lobe function tasks. Stepwise regression analysis revealed that relative amplitude and age represented the best predictors for cognitive performance (Stroop colour–word interference task, Trail Making Test A and B, semantic verbal fluency task), whereas psychopathology (Positive and Negative Syndrome Scale) did not significantly correlate with either cognitive performance levels or the quality of sleep–wake cycles. Conclusions Consolidated circadian rhythms and sleep may be a prerequisite for adequate cognitive functioning in individuals with schizophrenia

Autoantibodies Targeting AT₁- and ET_A-Receptors Link Endothelial Proliferation and Coagulation via Ets-1 Transcription Factor

Scleroderma renal crisis (SRC) is an acute life-threatening manifestation of systemic sclerosis (SSc) caused by obliterative vasculopathy and thrombotic microangiopathy. Evidence suggests a pathogenic role of immunoglobulin G (IgG) targeting G-protein coupled receptors (GPCR). We therefore dissected SRC-associated vascular obliteration and investigated the specific effects of patient-derived IgG directed against angiotensin II type 1 (AT1R) and endothelin-1 type A receptors (ETAR) on downstream signaling events and endothelial cell proliferation. SRC-IgG triggered endothelial cell proliferation via activation of the mitogen-activated protein kinase (MAPK) pathway and subsequent activation of the E26 transformation-specific-1 transcription factor (Ets-1). Either AT1R or ETAR receptor inhibitors/shRNA abrogated endothelial proliferation, confirming receptor activation and Ets-1 signaling involvement. Binding of Ets-1 to the tissue factor (TF) promoter exclusively induced TF. In addition, TF inhibition prevented endothelial cell proliferation. Thus, our data revealed a thus far unknown link between SRC-IgG-induced intracellular signaling, endothelial cell proliferation and active coagulation in the context of obliterative vasculopathy and SRC. Patients’ autoantibodies and their molecular effectors represent new therapeutic targets to address severe vascular complications in SSc